Abstract
Thioredoxin (Trx) is small ubiquitous protein that catalyzes thiol-disulfide reaction by two catalytic cysteine. Green sulfur bacterium Chlorobaculum tepidum TLS require strict anaerobic condition for growth and use various inorganic sulfur compounds as well as H2S as electron donors. To elucidate thioredoxin system in C. tepidum, we used thioredoxin mutant which one cysteine at the active site was substituted with serine. By immobilized cysteine mutant to resin, malate dehydrogenase (MDH) was captured as a possible target protein in C. tepidum. To further dissect this interaction, we used recombinant MDH that could be oxidized and rendered inactive. Inactivation was reversed by incubation with C. tepidum thioredoxin and dithiothreitol. Inactivation of MDH was found to result from formation of an intramolecular disulfide bond that was efficiently reduced by thioredoxin. These results demonstrate that the oxidized MDH is a preferable target protein of C. tepidum thioredoxin as suggested by thioredoxin affinity chromatography.
